c-erbB-2/HER-2/neu Antibody Staining Protocol for Immunohistochemistry

 

Description: c-erbB-2, second member (c-erbB- 2/HER-2/neu) of the c-erbB family is a receptor tyrosine kinase. It exhibits extracellular domains with two cysteine-rich sequences, and a cytoplasmic tyrosine kinase domain flanked by large hydrophilic tails that carry several tyrosine autophosphorylation sites. Approximately 25% of primary breast and ovarian tumors were found to overexpress the protein.

 

Primary Antibody

Name: c-erbB-2/HER-2/neu Ab-1 (21N) Antibody

Clone: Rabbit polyclonal

Supplier: Lab Vision

Catalog Number: RB-103-P

Dilution: 1:500 using IHC-TekTM Antibody Diluent (Cat# IW-1000 or IW-1001) to reduce background and unspecific staining and serum blocking step is NOT needed.

Incubation Time/Temp: 60 minutes/room temperature


Antigen Retrieval
Device: IHC-TekTM Epitope Retrieval Steamer Set (Cat# IW-1102)
Buffer/pH value: IHC-TekTM Epitope Retrieval Solution (Cat# IW-1100)
Heat/Cool Temperature: 95-100 ºC/room temperature
Heat/Cool Time: 20 minutes/20 minutes

Detection Methods
Standard Method: ABC Method or LSAB Method
Enhanced Method: Polymeric Methods

Chromogen Substrate
Reagent: DAB
Incubation Time/Temperature: 1-3 minutes/room temperature

Counterstain
Reagent: Mayer's Hematoxylin
Staining Time: 30 seconds

Results:
Staining Pattern: Membrane
Images: Search image

Additional Information:
Tissue Type: Breast carcinoma
Fixation: Formalin fixed paraffin sections
Positive Control: Breast carcinoma
Negative Control: Omit primary antibody, isotype control or absorption control
Blocking: 2-5% normal serum to reduce unspecific background staining; 0.5-3% H2O2 to block endogenous peroxidase activity; avidin/biotin to block endogenous biotin activity if necessary

 

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